Project Abstract The many complex behaviors that the human nervous system must perform rely on the existence of precise structural connections between neurons. During neural development, these precise connections are established by an initial period of axon growth and indiscriminant synapse formation, followed by a later period of activity-dependent synapse refinement, resulting in the formation of the mature circuit. Activity-dependent synapse refinement is therefore essential for the development of a healthy brain. Furthermore, abnormalities in these processes are implicated in the pathophysiological mechanisms of neurodevelopmental diseases. Work from the Shatz lab over the last several decades has sought to understand the molecular cues that link changes in neural activity to changes in synapses and circuits. This research uncovered a surprising role for the major histocompatibility class I (MHCI) molecules as important proteins in activity-dependent synaptic refinement. This was surprising because the MHCI molecules were best known for their role in the immune system, where they assist in defense against viruses and intracellular abnormalities. Research from the Shatz lab and others has revealed that in neurons, MHCI molecules are regulated by neuronal activity, and are required for the synapse elimination process that refines initial connections in the developing visual system into the full adult circuit. In addition, large-scale genome-wide association studies have identified polymorphisms in the MHCIs locus as strongly associated with several neuropsychiatric disorders, particularly schizophrenia. Post-mortem studies reveal that schizophrenic patients have fewer synapses than controls in some brain regions, further suggesting that schizophrenia may be a disorder of excess synapse elimination. However, until now, the bulk of the work linking MHCI and activity-dependent synapse elimination has been confined to model organisms. The role of human MHCIs in activity-dependent plasticity and synapse elimination has not previously been studied, and this represents a vital step forward in translating these discoveries further into a clinical context. The central goal of this proposal is to use histological, biochemical, electrophysiological, and genetic tools to determine the role of human MHCI molecules in activity-dependent synapse refinement. Here, I present preliminary work that establishes human brain organoids, 3D neuronal structures derived from induced pluripotent stem cells, as a feasible platform for studying MHCI and activity-dependent synaptic changes in human neurons. I will utilize these organoids to test the hypothesis that MHCI molecules can be found in human neurons, at synapses, where they function as components of an activity-dependent synaptic refinement mechanism. In the long-term, this research will provide a necessary link between previous mouse studies and human biology, opening the door for direct study of the molecular underpinnings of human neurodevelopment and neurodevelopmental disorders.